Ultrasound system for shearing cellular material in a microplate

ABSTRACT

Disclosed embodiments include illustrative piezoelectric element array assemblies, methods of fabricating a piezoelectric element array assembly, and systems and methods for shearing cellular material. Given by way of non-limiting example, an illustrative piezoelectric element array assembly includes at least one piezoelectric element configured to produce ultrasound energy responsive to amplified driving pulses. A lens layer is bonded to the at least one piezoelectric element. The lens layer has a plurality of lenses formed therein that are configured to focus ultrasound energy created by single ones of the at least one piezoelectric element into a plurality of wells of a microplate disposable in ultrasonic communication with the lens layer, wherein more than one of the plurality of lenses overlie single ones of the at least one piezoelectric element.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application is related to, and claims the benefit ofpriority of the filing date of, U.S. Provisional Patent Application No.62/448,857 filed Jan. 20, 2017, which is herein incorporated byreference in its entirety. The present application also relates toimprovements to subject matter disclosed in PCT Application No.PCT/US2015/040444 filed Jul. 14, 2015, which is herein incorporated byreference in its entirety.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

Disclosed subject matter was made with government support under grantnos. 1 R21 GM 111439-01 and 1 R33 CA 191135-01 awarded by the NationalInstitutes of Health. The government has certain rights in the disclosedsubject matter.

FIELD

The present disclosure is related to ultrasound systems for shearingcellular material.

BACKGROUND

The statements in this section merely provide background informationrelated to the present disclosure and may not constitute prior art.

Sample preparation is one of the preliminary steps that is performedbefore biological samples are analyzed. Sample preparation ofteninvolves the breakdown of the material into cellular or subcellularfragments. One particular application is the breaking up (or shearing)of DNA or Chromatin into smaller fragments. Ultrasound is one knownmethod of breaking down material. In some prior art devices, biologicalsamples are placed into a test tube that is put into a liquid bath andsubjected to high intensity ultrasound waves—similar to a jewelrycleaner, but with much higher power levels. To avoid an uneven exposureof the sample, the test tube is moved around within the variableultrasound field as it is processed. While this approach does work, itis limited to processing a single (or a few) test tube sample(s) at atime, thereby resulting in long processing times. Moreover, non-uniformultrasound fields may create hot and cold spots that require that thetest tube be moved around to get consistent shearing.

To increase the throughput of cellular processing, some currently knownsystems analyze cellular samples in microplates. As will be appreciatedby those skilled in the art, a microplate is a tray that contains anarray of wells in which samples can be placed for analysis. Advantagesof using microplates include processing such trays with automatedequipment and processing multiple samples at the same time withoutmoving the samples from one vessel to another.

One currently known system for shearing cellular samples in a microplateuses ultrasonically vibrating pins that extend into the wells. However,this can lead to cross contamination between the various wells andrequires extensive cleaning of the pins. It is also not very useful fortissue samples. Furthermore, the quality of the results depends greatlyon the exact position of the tips in the sample.

Another currently known approach uses a large ultrasound transducer thatis positioned below a single well and focuses the energy within thewell. The focused ultrasound energy creates cavitation in the samplematerial that is in the well but only one well is processed at a time.For a 96 element microplate, the processing time to shear all thesamples can exceed several hours during which some samples may degrade.

Another currently known approach uses a device that processes a columnor row of 8 or 12 DNA samples—but it cannot process chromatin. Thisdevice is a subset of 96 wells—not the entire microplate.

Another currently known approach to processing cellular material in amicroplate is to place a single ultrasound transducer below each well.See, for example, U.S. Pat. No. 6,699,711 to Hahn et al. (“Hahn”).However, when trying to experiment with the system described in the Hahnpatent for use in analyzing biological materials including DNA andchromatin, it was found that the system was ineffective in shearingchromatin without causing the transducers to break.

Another currently known approach uses a large ultrasonic horn in whichthe entire microplate is processed at once. However, the wells are notprocessed evenly because the system operates at low kHz frequencies andthere are hot/cold spots that result in uneven processing.

SUMMARY

Disclosed embodiments include illustrative piezoelectric element arrayassemblies, methods of fabricating a piezoelectric element arrayassembly, and systems and methods for shearing cellular material.

In an embodiment, an illustrative piezoelectric element array assemblyis provided. The piezoelectric element array assembly includes at leastone piezoelectric element configured to produce ultrasound energyresponsive to amplified driving pulses. A lens layer is bonded to the atleast one piezoelectric element. The lens layer has a plurality oflenses formed therein that are configured to focus ultrasound energycreated by single ones of the at least one piezoelectric element into aplurality of wells of a microplate disposable in ultrasoniccommunication with the lens layer, wherein more than one of theplurality of lenses overlie single ones of the at least onepiezoelectric element.

In another embodiment, an illustrative method of fabricating apiezoelectric element array assembly is provided. The method includes:providing at least one piezoelectric element configured to produceultrasound energy responsive to amplified driving pulses; and bonding alens layer to the at least one piezoelectric element, the lens layerhaving a plurality of lenses formed therein that are configured to focusultrasound energy created by single ones of the at least onepiezoelectric element into a plurality of wells of a microplatedisposable in ultrasonic communication with the lens layer, wherein morethan one of the plurality of lenses overlie single ones of the at leastone piezoelectric element.

In another embodiment, an illustrative system for shearing cellularmaterial is provided. The system includes a signal generator configuredto generate ultrasound driving pulses. An amplifier is electricallycoupled to the signal generator and configured to amplify the ultrasounddriving pulses. A piezoelectric element array includes at least onepiezoelectric element configured to produce ultrasound energy responsiveto amplified driving pulses; and a plurality of lenses, wherein morethan one of the plurality of lenses overlie single ones of the at leastone piezoelectric element and wherein single ones of the plurality oflenses are configured to focus ultrasound energy into single ones of aplurality of wells of a microplate.

In another embodiment, an illustrative method includes: generatingultrasound driving pulses; amplifying the ultrasound driving pulses;producing ultrasound energy with at least one piezoelectric elementresponsive to the amplified driving pulses; and focusing the ultrasoundenergy created by single ones of the at least one piezoelectric elementinto a plurality of wells of a microplate by a plurality of lenses,wherein more than one of the plurality of lenses overlie single ones ofthe at least one piezoelectric element and wherein single ones of theplurality of lenses are ultrasonically coupled to single ones of theplurality of wells.

In another embodiment, another illustrative system for shearing cellularmaterial is provided. The system includes a computer processorconfigured to generate timing signals. A signal generator is configuredto generate ultrasound driving pulses responsive to the timing signals.An amplifier is electrically coupled to the signal generator and isconfigured to amplify the ultrasound driving pulses. A plurality ofpiezoelectric elements is arranged in an array of rows and columns. Thepiezoelectric elements are configured to produce ultrasound energyresponsive to amplified driving pulses. The timing signals are generatedsuch that adjacent ones of the plurality of piezoelectric elements arenot energized by at least amplified driving pulses chosen fromsimultaneous driving pulses and temporally sequential driving pulses.More than one of a plurality of lenses overlie single ones of theplurality of piezoelectric elements and single ones of the plurality oflenses are configured to focus ultrasound energy into single ones of aplurality of wells of a microplate.

In another embodiment, another illustrative system for shearing cellularmaterial is provided. The system includes a housing. A signal generatoris disposed in the housing and is configured to generate ultrasounddriving pulses. An amplifier is disposed in the housing and iselectrically coupled to the signal generator, and the amplifier isconfigured to amplify the ultrasound driving pulses. A piezoelectricelement array is disposed in the housing. The piezoelectric elementarray includes at least one piezoelectric element configured to produceultrasound energy responsive to amplified driving pulses. A plurality oflenses are configured to focus ultrasound energy into a plurality ofwells of a microplate. A fluidics system is configured to flow therein atransducer fluid. A seal is disposed on the housing. The seal isconfigured to receive a microplate in sealing engagement thereon suchthat the piezoelectric element array, the housing, and a microplatereceived in sealing engagement on the seal define a chamber in hydrauliccommunication with the fluidics system and configured to contain thereintransducer fluid.

In an another embodiment, an illustrative method of shearing cellularmaterial is provided. The method includes: placing a microplate withcellular material disposed in a plurality of wells defined therein on aseal disposed on a housing; clamping the microplate on the seal insealing engagement therewith; flowing transducer fluid in a fluidicssystem disposed in the housing such that transducer fluid is placed inhydraulic communication with a plurality of lenses; energizing an arrayof piezoelectric elements to produce ultrasound energy; and focusingultrasound energy in the plurality of wells with a plurality of lensessuch that cavitation is induced in the cellular material disposed in theplurality of wells.

Further features, advantages, and areas of applicability will becomeapparent from the description provided herein. It should be understoodthat the description and specific examples are intended for purposes ofillustration only and are not intended to limit the scope of the presentdisclosure.

BRIEF DESCRIPTION OF THE DRAWINGS

The drawings described herein are for illustration purposes only and arenot intended to limit the scope of the present disclosure in any way.The components in the figures are not necessarily to scale, withemphasis instead being placed upon illustrating the principles of thedisclosed embodiments. In the drawings:

FIG. 1A is an exploded perspective view of an illustrative piezoelectricelement array assembly.

FIG. 1B is an exploded perspective view of another illustrativepiezoelectric element array assembly.

FIG. 1C is an exploded perspective view of another illustrativepiezoelectric element array assembly.

FIG. 1D is an exploded perspective view of another illustrativepiezoelectric element array assembly.

FIG. 1E is an exploded perspective view of another illustrativepiezoelectric element array assembly.

FIG. 1F is an exploded perspective view of another illustrativepiezoelectric element array assembly.

FIG. 2A is a perspective view of an illustrative array of piezoelectricelement array assemblies.

FIG. 2B is a side plan view in partial cutaway of a portion of the arrayof piezoelectric element array assemblies of FIG. 2A.

FIG. 2C is a top plan view of the array of piezoelectric element arrayassemblies of FIG. 2A.

FIG. 3 is an exploded perspective view in partial schematic form of anillustrative system for shearing cellular material.

FIG. 4A is a perspective view in partial schematic form of anillustrative piezoelectric element underlying lenses formed in wells ofan illustrative microplate.

FIGS. 4B and 4D are end plan views in partial schematic form of an arrayof the piezoelectric elements of FIG. 4A underlying lenses formed inwells of an illustrative microplate.

FIG. 4C is a perspective view in partial cutaway of details of a well ofa microplate.

FIG. 4E is an exploded perspective view in partial schematic form ofanother illustrative system for shearing cellular material.

FIG. 5 is a block diagram in partial schematic form of anotherillustrative system for shearing cellular material.

FIG. 6 is a piping diagram in partial schematic form of details of afluidics system of the system of FIG. 5.

FIG. 7A is a perspective view of an illustrative system for shearingcellular material.

FIGS. 7B-7D are perspective views in partial cutaway of details of thesystem of FIG. 7A.

FIG. 7E is a front plan view in partial cutaway of details of the systemof FIG. 7A.

FIG. 7F is a front plan view in partial cutaway of details of anotherembodiment of the system of FIG. 7A.

FIGS. 8A-8D illustrate details of waveform timing of the system of FIG.5.

FIG. 8E illustrates further details of the system of FIG. 5.

DETAILED DESCRIPTION

The following description is merely illustrative in nature and is notintended to limit the present disclosure, application, or uses.

As will be discussed in further detail below, disclosed embodimentsinclude illustrative piezoelectric element array assemblies, methods offabricating a piezoelectric element array assembly, and systems andmethods for shearing cellular material. Given by way of overview, invarious embodiments a sufficient amount of ultrasound energy is appliedsimultaneously to a number of samples in order to cause inertialcavitation to occur in the samples, thereby causing some shearing ofmolecular bonds of DNA and chromatin in the samples.

As discussed above, one of the difficulties of using ultrasound to shearthe cellular material is that some currently known transducer elementscan crack when driven to a level that is sufficient to cause cavitation.Various embodiments of subject matter disclosed herein relate toimprovements to transducer design that improve the durability of thetransducer elements.

Still by way of overview, various embodiments of subject matterdisclosed herein relate to illustrative piezoelectric element arrayassemblies that can focus ultrasound energy from a single piezoelectricelement to more than one well of a microplate and to illustrativemethods for fabrication of such piezoelectric element array assemblies.Various embodiments of subject matter disclosed herein also relate tosystems and methods for shearing cellular material in which suchpiezoelectric element array assemblies may be employed. Variousembodiments of subject matter disclosed herein also relate to systemsand methods for shearing cellular material in which improvements havebeen made to currently known aspects related to ultrasonically couplingpiezoelectric element arrays to microplates with transducer fluid.

Now that an overview has been provided, details will be set forth belowby way of non-limiting examples and not of limitation.

Referring now to FIG. 1A, an illustrative piezoelectric element arrayassembly 10 can focus ultrasound energy from a single piezoelectricelement 12 to more than one well of a microplate (not shown). In variousembodiments, the piezoelectric element array assembly 10 includes atleast one piezoelectric element 12 configured to produce ultrasoundenergy responsive to amplified driving pulses. A lens layer 14 is bondedto the piezoelectric element 12. The lens layer 14 has lenses 16 formedtherein. Each lens 16 is configured to focus ultrasound energy into onewell of a microplate (not shown in FIG. 1A) that is disposed inultrasonic communication with the lens layer 12. As shown in FIG. 1A,more than one lens 16 overlies the single piezoelectric element 12. Assuch, the piezoelectric element array assembly 10 can focus ultrasoundenergy from a single piezoelectric element 12 to more than one well of amicroplate (not shown in FIG. 1A)—because more than one lens 16 overliesthe single piezoelectric element 12.

Still referring to FIG. 1A, in various embodiments the piezoelectricelement 12 may include a strip of piezoelectric substrate material suchas lead zirconate titanate (Pb[Zr(x)Ti(1-x)]O3) (“PZT”) that is coatedor plated on both sides with a conductor such as aluminum, gold, copper,or the like. The piezoelectric element 12 has width, length, andthickness dimensions that are selected to prevent cracking when drivenwith a voltage signal that is sufficient to cause cavitation in a wellof a microplate.

In various embodiments each piezoelectric element 12 may have a widthequal to the width of a single well of a microplate and a lengthselected to underlie more than one well of a microplate. Accordingly, itwill be appreciated that more than one lens 16 may overlie any singlepiezoelectric element 12. Thus, the piezoelectric element array assembly10 can focus ultrasound energy from a single piezoelectric element 12 tomore than one well of a microplate (not shown in FIG. 1A) because morethan one lens 16 overlies the single piezoelectric element 12.

As discussed above, in some embodiments each piezoelectric element 12may have a length selected to underlie more than one well of amicroplate. In some embodiments and as shown in FIG. 1A, thepiezoelectric element 12 may have a length selected to underlie fourwells of a microplate and a width equal to the width of a single well.As such, four of the lenses 16 overlie the single piezoelectric element12. Using conventional microplate dimensions, the length and width ofthe piezoelectric element 12 as shown in FIG. 1A is approximately 36×9mm.

In various embodiments the lens layer 14 has concave lenses 16 formed inan upper surface thereof. The lenses 16 operate to focus the ultrasonicplane waves created by the piezoelectric element 12 as the piezoelectricelement 12 is excited by a driving voltage. The lenses 16 are shaped tofocus the plane waves into a well of a microplate (not shown) that ispositioned above the lens layer 14.

It will be appreciated that in other embodiments piezoelectric elements12 of other sizes could be used and any number of lenses 16 may overliethe piezoelectric element 12 as desired for a particular application.For example and referring to FIG. 1B, in some embodiments thepiezoelectric element 12 may have a length selected to underlie twowells of a microplate and a width equal to the width of a single well.As such, two of the lenses 16 overlie the single piezoelectric element12. As another example and referring to FIG. 1C, in some embodiments thepiezoelectric element 12 may have a length selected to underlie sixwells of a microplate and a width equal to the width of a single well.As such, six of the lenses 16 overlie the single piezoelectric element12. As another example and referring to FIG. 1D, in some embodiments thepiezoelectric element 12 may have a length selected to underlie eightwells of a microplate and a width equal to the width of a single well.As such, eight of the lenses 16 overlie the single piezoelectric element12. As another example and referring to FIG. 1E, in some embodiments thepiezoelectric element 12 may have a length selected to underlie twelvewells of a microplate and a width equal to the width of a single well.As such, twelve of the lenses 16 overlie the single piezoelectricelement 12. However, no limitation other to the number of lenses 16which may overlie the single piezoelectric element 12 is intended and isnot to be inferred—other than more than one lens 16 overlies the singlepiezoelectric element 12. Moreover, no limitation is intended to thewidth of the piezoelectric elements 12, which may have any width asdesired for a particular application, such as arrays of 2×2, 2×4, 3×4,and the like.

It will be appreciated that, in each embodiment, the piezoelectricelement 12 is sized to deliver ultrasound energy to two or more wells inthe microplate so that the stresses created in the strip are spread outover an area that is larger than the area of a single well. In oneembodiment, the thickness of the piezoelectric element 12 is selected toproduce ultrasound energy at a frequency selected between 500 KHz and 2MHz.

Referring now to FIG. 1F and depending on the impedance of the lenslayer 14, in some embodiments an optional matching layer 18 may bepositioned between the piezoelectric element 12 and the lens layer 14.In various embodiments the matching layer 18 has a thickness of ¼wavelength at the operating frequency of the system and is bonded to theupper surface of the piezoelectric element 12 and the bottom of the lenslayer 14 with an adhesive, such as without limitation epoxy. It will beappreciated that the optional matching layer 18 may be used inconjunction with piezoelectric elements 12 of any size, such as thosediscussed above.

In various embodiments, the matching layer 18 may be omitted. Forexample, in some embodiments the lens layer 14 is made of a materialhaving an acoustic impedance that is between the impedance of thepiezoelectric element 12 and a coupling fluid (not shown), such aswithout limitation water or gel, that is placed in ultrasoniccommunication between the lens layer 14 and the wells of a microplate.In such embodiments, the matching layer 18 suitably is omitted. In someembodiments the lens layer 14 may be made of a suitable material such asgraphite, fluorphlogopite mica in a borosilicate glass matrix, and thelike.

Referring now to FIGS. 2A-2C, embodiments of piezoelectric element arrayassemblies 10 a, 10 b, 10 c, 10 d . . . 101 are arranged in an array 20of rows and columns to deliver ultrasound energy to all the wells of aninety-six (96) well microplate (not shown). It will be appreciated thatin various embodiments a larger or smaller number of piezoelectricelement array assemblies 10 could be used to accommodate different sizedmicroplates. It will also be appreciated that, while the piezoelectricelement array assemblies 10 a, 10 b, 10 c . . . 101 are shown as beingsized such that four (4) lenses 16 overlie each piezoelectric elementarray assembly 10, in various embodiments the piezoelectric elementarray assemblies 10 may be sized as desired such that two or more lenses16 overlie each piezoelectric element array assembly 10 (as shown inFIGS. 1A-1E). In various embodiments the piezoelectric element arrayassemblies 10 are preferably held in a frame (not shown) that maintainsthe arrangement of the piezoelectric element array assemblies 10 belowthe microplate (not shown). In some embodiments, the size of thepiezoelectric elements 12 may be slightly larger than the size of thelens layer 14 in order to allow a wrap-around electrode (not shown) tobe used to connect electrical wires to the piezoelectric elements 12.

Referring to FIG. 3, in various embodiments and as an introduction toillustrative system environments in which cellular material is shearedwith ultrasound energy, a signal generator 22 produces driving signalsthat are amplified by an amplifier 24 and each piezoelectric element 12is electrically connected to the amplifier 24. In various embodiments,the driving signals may be waveforms such as short bursts of pulses(such as without limitation 10-50 microseconds or so) that are amplifiedto a sufficient voltage level (such as approximately 400 V) so that theacoustic energy produced is sufficient to cause inertial cavitation inthe cellular material in a well. In some embodiments, the bursts arespaced in time to reduce heating the sample and the coupling fluid.

In some embodiments each piezoelectric element 12 may be electricallyconnected to the amplifier 24 individually. In some other embodimentspiezoelectric elements 12 may be electrically connected in parallel. Insome such embodiments, adjacent piezoelectric elements 12 may beelectrically connected in parallel to the amplifier 24. In other suchembodiments, the parallel electrical connections may be spaced apartover the pattern of piezoelectric elements 12 so that, when energized,adjacent piezoelectric elements 12 are not vibrating.

Referring additionally to FIGS. 4A-4E, in other embodiments the lenses16 may be formed in wells of a microplate 210 instead of being formed ina lens layer that is bonded to the piezoelectric elements 12. As shownin FIGS. 4A, 4B, and 4D, in such embodiments the piezoelectric elementarray 10 includes the piezoelectric elements 12. Each one of the lenses16 is formed in single ones of wells 210 of a microplate (not shown).

As shown in FIG. 4C, in such embodiments the bottom of the well 210 canbe molded as the lens 16 to focus ultrasound energy into the well 210and, as such, may not require a separate focusing lens 16 between thepiezoelectric element 12 and the bottom of the microplate (as shown inFIGS. 1A-1F, 2A-2C, and 3). As shown in FIG. 4C, the well 210 has thelens 16 integrally formed therein. As shown in FIGS. 4C and 4D, couplingmaterial—that is, transducer fluid 117 (described below)—is positionedbetween the bottom of the well 210 and the piezoelectric element 12. Invarious embodiments, the bottom of the well 210 may have a concave shapeto act as a lens 16 that focuses ultrasound energy into an interiorportion of the well 210. It will be appreciated that the well 210 may beinjection-molded to form the lens 16 in its desired shape and focus theultrasound energy into the desired portion of the well 210. It will alsobe appreciated that cellular material in the well 210 is sheared due toinertial cavitation occurring in the well 210.

Referring now to FIG. 4E, in some embodiments the signal generator 22produces driving signals that are amplified by the amplifier 24 and eachpiezoelectric element 12 is electrically connected to the amplifier 24.In such embodiments each one of the lenses 16 is formed in single onesof wells 210 of a microplate (not shown).

Now that an introduction to illustrative system environments has beenset forth, embodiments of various illustrative system environments willbe described by way of non-limiting examples.

Referring now to FIG. 5, in various embodiments an illustrative system100 is provided for shearing cellular material. In some of theseembodiments, the system 100 includes the various embodiments of thepiezoelectric element array assemblies 10 in which more than one lens 16overlies a single piezoelectric element 12 (FIGS. 1A-1F, 2A-2C, and4A-4D). Other embodiments of the system 100 may use any suitablepiezoelectric element array assemblies as desired, such as thepiezoelectric element array assemblies 10 in which more than one lens 16overlies a single piezoelectric element 12 and also such aspiezoelectric element array assemblies in which only one lens overlies asingle piezoelectric element.

Still referring to FIG. 5 and given by way of overview, in someembodiments the system 100 includes the various embodiments of thepiezoelectric element array assemblies 10 in which more than one lens 16overlies a single piezoelectric element 12. In such embodiments, thesystem 100 includes a computer processor 102 configured to generatetiming signals 104. A signal generator 106 is configured to generateultrasound driving pulses 108 responsive to the timing signals 104. Anamplifier 110 is electrically coupled to the signal generator 106 and isconfigured to amplify the ultrasound driving pulses 108. Piezoelectricelements 12 are arranged in an array 112 of rows and columns (such aswithout limitation the array 20 shown in FIGS. 2A-2C or the array ofpiezoelectric elements 10 shown in FIGS. 4B and 4D). It will beappreciated that in various embodiments the array 112 may include 1×Narrays and in some other embodiments may include >1×N arrays (such as,without limitation, 2×2, 2×4, 3×3, and the like). The piezoelectricelements 12 are configured to produce ultrasound energy responsive toamplified driving pulses 114. The timing signals 104 are generated suchthat adjacent piezoelectric elements 12 are not energized bysimultaneous amplified driving pulses 114 and/or temporally sequentialamplified driving pulses 114. In some embodiments, a lens layer 14(FIGS. 1A-1F and 2A-2C) is bonded to the piezoelectric elements 12(FIGS. 1A-1F and 2A-2C). The lens layer 14 has lenses 16 (FIGS. 1A-1Fand 2A-2C) formed therein. In some other embodiments (FIGS. 4A-4D), thelenses 16 (FIGS. 4A-4D) are formed in the wells 210 (FIGS. 4A-4D) of amicroplate. More than one of the lenses 16 (FIGS. 1A-1F, 2A-2C, and4A-4D) overlie single piezoelectric elements 12 (FIGS. 1A-1F, 2A-2C, and4A-4D) and single lenses 16 (FIGS. 1A-1F, 2A-2C, and 4A-4D) areconfigured to focus ultrasound energy into single wells of a microplate(not shown).

Still referring to FIG. 5 and still given by way of overview, in someembodiments the system 100 may use any suitable piezoelectric elementarray assemblies as desired, such as the piezoelectric element arrayassemblies 10 (FIGS. 1A-1F, 2A-2C, and 4A-4D) in which more than onelens 16 overlies a single piezoelectric element 12 and also such aspiezoelectric element array assemblies in which only one lens overlies asingle piezoelectric element. In such embodiments, the system 100includes a housing (not shown in FIG. 5). The signal generator 106 isdisposed in the housing and is configured to generate the ultrasounddriving pulses 108. The amplifier 110 is disposed in the housing and iselectrically coupled to the signal generator 106, and the amplifier 110is configured to amplify the ultrasound driving pulses. Thepiezoelectric element array 112 is disposed in the housing. Thepiezoelectric element array 112 includes at least one piezoelectricelement configured to produce ultrasound energy responsive to theamplified driving pulses 114. Lenses are configured to focus ultrasoundenergy into wells of a microplate. A fluidics system 115 is configuredto flow therein a transducer fluid 117 that ultrasonically couples thepiezoelectric elements and a microplate. In various embodiments, thetransducer fluid 117 may include any suitable fluid, such as withoutlimitation a solution of a surfactant and water. It will be appreciatedthat, in some embodiments, a surfactant can wet the bottom of themicroplate so that bubbles do not form there and possibly block theultrasound energy from entering the sample contained in the well. A seal(not shown in FIG. 5) is disposed on the housing. The seal is configuredto receive a microplate in sealing engagement thereon such that thepiezoelectric elements, the housing, and a microplate received insealing engagement on the seal define a chamber in hydrauliccommunication with the fluidics system 115 and configured to containtherein the transducer fluid 117.

Now that an overview of various embodiments of the system 100 have beenset forth, details will be explained below by way of illustrative,non-limiting examples. Functional details will be addressed first,followed by mechanical details.

Still referring to FIG. 5, in various embodiments a user interface 116is electrically coupled to the computer processor 102. The userinterface may include a graphical user interface, such as any suitable,commercially available touchscreen. The user interface 116 displaysinformation to a user and accepts the user response(s) to the displayedinformation, thereby permitting a user to set up and control the system100. In various embodiments, via the user interface 116 a user may enterinformation such as without limitation: process selection (such asChromatin, DNA, or Service protocol); column(s) within the array 112selected for processing (such as any or all of the columns); processingtime (for all selected columns); start process (for all selectedcolumns); pause process (all columns); power levels (for all or a subsetof wells); pulse parameters (burst length, column-to-column cyclingtime, PRF, duty cycle, or a combination of parameters); and Restart orCancel process (while in a paused state). In various embodiments, theuser interface 116 may display to a user parameters such as withoutlimitation: device state (which is inherent in the displayedinformation); process selected; column(s) selected; processing timeselected; process time elapsed/remaining (progress indicator) duringprocessing; and process complete indication.

In various embodiments the computer processor 102, via the userinterface 116, displays device states and options to the user andreceives user inputs. It will be appreciated that the computer processor102 is disposed in a housing (discussed below). That is, the computerprocessor 102 is integrated into the system 100 instead of being astand-alone unit, such as a laptop or desktop computer, that residesoutside the physical boundaries of the system 100. The computerprocessor 102 also configures the system 100 for operation, controls theoverall process timing, initiates processes, pauses processes, resumesprocesses, and monitors state of the system 100. The computer processor102 suitably is any commercially available computer processor. Given byway of non-limiting example, in various embodiments the computerprocessor may be a Linux-based computer processor such as, for example,Raspberry Pi (a single board computer processor). In variousembodiments, the computer processor 102 controls the following processparameters: which column(s) is(are) active; processing time; pulseparameters; and the functions ON (enables output waveform), PAUSE(pauses output waveform), RESUME (resumes output waveform timing), andCANCEL cancels process and resets timing and returns to menu state). Invarious embodiments the computer processor 102 receives the followinginputs: user input signals 118 from the user interface 116; a signal 120from the signal generator 106 for heartbeat (to indicate normaloperation of the signal generator 106)/watchdog (to prevent the signalgenerator 106 from elapsing or timing out; a safety interlock statesignal 122 (discussed below); a transducer fluid level monitoring signal124 (discussed below); and a transducer fluid temperature monitoringsignal 126 (discussed below). In various embodiments, the computerprocessor 102 provides the following outputs: a display interface signal128 supplied to the user interface 116; a column enable signal 130 foractivating selected columns; a waveform selection (that is, the timing)signal 104; an Output ON or Output OFF signal 132; a reset waveformsequencer controller timing signal 134 that resets the timing of thesignal generator 106; pulse parameters; and a fluidics control signal136 (discussed below).

In various embodiments, the signal generator 106 generates waveforms(that is, the driving pulses 108) to drive the amplifier 110. The signalgenerator 106 also gates the driving pulses 108 to specific column(s) ofthe array 112 based on the column selected and the safety interlockbeing engaged (via the safety interlock state signal 122). In variousembodiments the signal generator 106 suitably may be a commercialoff-the-shelf field-programmable gate array (FPGA)-based module. In someembodiments the signal generator 106 may include multiple FPGA modules137 (depending on FPGA capacity). In embodiments in which the signalgenerator 106 includes more than one FPGA module, then the number ofFPGA modules may equal the number of amplifier modules (discussed below)to simplify the architecture. In various embodiments the signalgenerator receives the following inputs: the column enable signal 130for activating selected columns; the waveform selection (that is, thetiming) signal 104; the Output ON or Output OFF signal 132; the resetwaveform sequencer controller timing signal 134 that resets the timingof the signal generator 106; and the safety interlock state signal 122.In various embodiments the signal generator provides the followingoutputs: waveforms (that is, the driving pulses 108); theheartbeat/watchdog signal 120; and a waveform counter signal 138 thatindicates current output elapsed time.

In various embodiments the amplifier 110 incorporates a suitable numberof pulser modules 140 as determined by the number of piezoelectricelements in the array 112 and by the number of channels per pulsermodule 140. The amplifier 110 generates transducer drive waveforms (thatis, the amplified driving pulses 114) with timing equal to therespective input column waveform (that is, the driving pulses 108). Theamplifier 110 may also provide a matching network to provide forintegrated tuning—that is, low pass filtering to limit radiatedelectromagnetic interference (EMI) and/or other filtering or impedancematching elements, thereby helping to result in increased power transferfor the transducers of the array 112. The amplifier 110 also integratesvoltage waveforms (that is, the amplified driving pulses 114) at theoutput of the amplifier 110. In various embodiments, the amplifier 110receives the following inputs: the differential waveform (that is, thedriving pulses 108) for the pulser channels; and a single high voltagesupply to bias the pulser modules 140. In various embodiments, theamplifier 110 provides the following outputs: the high power waveformfor the transducer columns (that is, the amplified driving pulses 114);and an analog voltage proportional to the integrated voltage to thetransducer column (that is, proportional to the amplified driving pulses114) that is provided to the computer processor 102 for monitoring andself-diagnostics purposes.

In various embodiments a monitor module 146 suitably receives thewatchdog/heartbeat signal 120, the transducer fluid level monitoringsignal 124, the transducer fluid temperature monitoring signal 126, thewaveform counter signal 138, the analog voltage 144, and voltages 148(from a power distribution module 150 that converts mains power intodirect current power for the components of the system 100) and providesthem to the computer processor 102. The monitor module 146 includessuitable circuitry and logic for performing self-diagnostics functionsfor the system 100.

Referring additionally to FIG. 6, in various embodiments the fluidicssystem 115 chills and circulates the transducer fluid 117 for couplingthe ultrasound energy from the piezoelectric elements and lenses to thewells of a microplate. It will be appreciated that all components of thefluidics system 115 are disposed within one housing (not shown in FIG. 6but discussed below)—as are all components of the system 100.

In various embodiments a reservoir 152 is configured to receive andstore transducer fluid 117. A suction port 154 of a pump 156 ishydraulically coupled to the reservoir 152 to receive transducer fluid117. The fluidics control signal 136 is supplied to control circuitry(not shown) of the pump 156, and state of flow in the fluidics system115 (that is, forward, off, or reverse) suitably is controlled by thefluidics control signal 136. A discharge port 158 of the pump 156 ishydraulically coupled to an inlet port 160 of a suitable filter 162,such as without limitation a particle filter. An outlet port 164 of thefilter 162 is hydraulically coupled to an inlet port 166 of a suitablechiller 168, such as without limitation a thermo-electric device like apeltier cooler. State of the chiller (that is, chiller on or chilleroff) suitably is controlled by the fluidics control signal 136. Anoutlet port 170 of the chiller 168 is hydraulically coupled to an inletport 172 of a well 174 that contains therein transducer fluid 117 forcoupling the ultrasound energy from the piezoelectric elements andlenses to the wells of a microplate 176. An outlet port 178 of the well174 is hydraulically coupled to the reservoir 152. A temperature probe180 provides the transducer fluid temperature monitoring signal 126 tothe monitor module 146 (FIG. 5) as monitored at an outlet of aprocessing tray 194. A fluid level probe 182 in the processing tray 194provides the transducer fluid level monitoring signal 124 to the monitormodule 146 (FIG. 5). The fluid level probe 182 is configured to monitorlevel of transducer fluid 117 in the processing tray 194 to help ensurethat a sufficient amount of transducer fluid 117 is contained in theprocessing tray 194 to ultrasonically couple the microplate 176 and thepiezoelectric elements. While flow impedance between the well 174 andthe reservoir 152 is maintained low, a seal (discussed below) betweenthe microplate 176 and the processing tray 194 can help to facilitatehigher flow rates of the transducer fluid 117.

It will be appreciated that, in some embodiments, debubbling anddegassing of the transducer fluid 117 is not required. It will also beappreciated that, in some other embodiments, debubbling and degassing ofthe transducer fluid 117 may be desired. In such embodiments in whichdebubbling and degassing of the transducer fluid 117 may be desired,optional debubbling and degassing components 184 may be interposed inthe fluidics system 115. As shown in FIG. 5, the optional debubbling anddegassing components 184 may include: a restrictor 186 interposedbetween the reservoir 152 and the suction port 154 of the pump 156; adebubbler 188 hydraulically coupled to the discharge port 158 of thepump 156; a degassing filter 190 interposed between the debubbler 188and the inlet port 160 of the filter 162; and a restrictor 192interposed between the debubbler 188 and the reservoir 152.

Mechanical/fluidics aspects will be discussed next, followed by adiscussion of functionality and operation of various embodiments of thesystem 100.

Referring additionally to FIGS. 7A-7E, in various embodiments of thesystem 100 all components of the system 100 are disposed in a housing196. The housing may be made of any suitable material as desired, suchas plastic, metal, or the like. In some embodiments the housing 196 mayfunction as a faraday shield. In some such embodiments the housing 196may be made of plastic and lined with either a continuous covering ofconductive material (not shown) or a mesh of conductive material (notshown). In such embodiments, an external electrical field causeselectric charges within the conductive material that lines the housing196 to be distributed such that the distributed electric charges cancelthe electric field's effect in the interior of the housing 196, therebyhelping to protect electronic components of the system 100 from externalradio frequency interference—and vice versa.

In various embodiments the housing includes a lid 198. In variousembodiments the lid 198 is configured to open and close, such as bybeing rotated upwardly and downwardly, respectively, about a hinged axisdisposed at an edge of the lid 198 toward a central part of the housing196. The lid 198 and an associated sunken portion 200 (FIGS. 7B and 7C)defined in the housing 196 are sized such that the microplate 176 can bereceived in the sunken portion 200 when the lid 198 is open. Closing thelid 198 (which means that a user has no access to the acoustic field)causes the safety interlock state signal 122 (FIGS. 5 and 6) to beactive. As a result, processing may be enabled. Opening the lid 198(which means that a user has access to the acoustic field) causes thesafety interlock state signal 122 (FIGS. 5 and 6) to be inactive. As aresult, processing is not enabled.

In various embodiments and as shown in FIGS. 7B and 7C, a seal 202 isdisposed on a top surface of the housing 196 within the sunken portion200. An opening (not shown) is defined in the top surface of the housing196 within the sunken portion 200 and the seal 202 surrounds theopening. The seal 202 is sized to receive thereon the microplate 176. Ahold-down frame 204 is placed on top of the microplate 176. Each of apair of clamps 206 urges a side of the hold down frame 204 onto themicroplate 176 in sealing engagement with the seal 202. Thus, thehousing 196, the seal 202, the microplate 176, the hold-down frame 204,and the clamps 206 cooperate to hydraulically seal the opening definedin the top surface of the housing 196 within the sunken portion 200.

In some embodiments, the hold down frame 204 and the clamps 206 may beintegrated into an underside of the lid 198. In such embodiments,closing the lid 198 applies the force entailed to cause the clamps 206to sealingly engage the housing 196, the seal 202, the microplate 176,and the hold-down frame 204. Conversely, opening the lid 198 causes theclamps 206 to disengage engage the housing 196, the seal 202, themicroplate 176, and the hold-down frame 204 from sealing engagement,thereby permitting removal of the microplate 176.

In various embodiments and as shown in FIG. 7D, the reservoir 152 issealed and filled with a lid 208 that is received on a top portion ofthe reservoir 152 (such as by being threadedly received or press fit).The lid is accessible in the sunken portion 200 of the housing 196. Insome embodiments, the reservoir 152 may include a level indicatordevice, such as without limitation an integrated sight glass/tube or thelike, to provide indication to a user regarding level of transducerfluid 117 in the reservoir 152. If desired, a sensor may provide a userwith knowledge about when to refill the reservoir 152.

In various embodiments and as shown in FIGS. 7E and 7F, the seal 202holds down the microplate 176 and the clamps 206 hold down the seal 202.Transducer fluid 117 in a gap 207 between the lenses 16 and themicroplate 176 ultrasonically couples the piezoelectric elements 12 withwells 210 of the microplate 176. The array 112 is attached to a baseframe 212 that is, in turn, attached to the housing 196. A printedcircuit board interface 214 electrically couples the piezoelectricelements 12 to cables (not shown) that electrically conduct theamplified driving pulses 114 from the amplifier 110.

In various embodiments of the system 100, no transducer fluid 117 is inthe gap 207. After the microplate 176 is sealingly engaged by the clamps206 to the seal 202 and the lid 198 is shut, transducer fluid 117 ispumped into the gap 207 from the reservoir 152 by the pump 156. Placingthe microplate 176 onto a bath-like structure with no transducer fluidinitially disposed therein and then filling the gap 207 with transducerfluid 117 only after the microplate 176 has been placed thereon flies inthe face of conventional systems in which a microplate is placed into abath of transducer fluid. As a result, embodiments of the system 100help provide more secure placement of the microplate 176 than inconventional systems and can help to reduce the possibility oftransducer fluid 117 getting into the wells 210 of the microplate 176,thereby helping to reduce the possibility of contaminating contents ofthe wells 210 with transducer fluid 117.

When the lid 198 is shut, the safety interlock is engaged and the safetyinterlock state is active (via the safety interlock state signal 122).The microplate 176 is sealingly engaged with the housing 196. Processingof samples can proceed as discussed below.

Referring now to FIGS. 1A-1F, 2A-2C, 3, 4A-4D, 5-6, 7A-7F, and 8A-8E,functionality and operation of various embodiments of the system 100will be explained by examples provided by way of illustration and not oflimitation.

Regarding control of embodiments of the system 100, the computerprocessor 102 is the center of control for the system 100. The computerprocessor 102 outputs display to the user interface 116 and reads userinput that is input via the user interface 116. In addition, thecomputer processor 102 sets up the signal generator 106 with respect towhich processing regimen is to be run. The computer processor 102directly controls which channels are active, the overall processingtime, starting the process, and stopping the process.

Waveform timing for driving the piezoelectric elements 12 output by thesignal generator as the driving pulses 108. In some embodiments of thesystem 100 with a ninety-six well microplate 176, four lenses 16 overlieeach piezoelectric element 12, and the array 112 includes twenty-fourpiezoelectric element array assemblies 10. In some embodiments, thenumber of pulser modules 140 may be equal to the number of piezoelectricelements 10, and in some other embodiments the number of pulser modules140 may be equal to half the number of piezoelectric elements 10.

As discussed above, for the waveform (that is, the driving pulses 108)to be output, the lid 198 must be closed. When the lid 198 is open, boththe computer processor 102 and the signal generator 106 do not allow theoutput (that is, the driving pulses 108) to be driven. This is becausethe safety interlock state signal 122 is input to both the computerprocessor 102 and the signal generator 106. In addition, the computerprocessor 102 monitors both the transducer fluid temperature (via thetransducer fluid temperature monitoring signal 126) and the transducerfluid level (via the transducer fluid level monitoring signal 124). Ifthe temperature of the transducer fluid 117 is above an acceptable levelor the transducer fluid 117 does not fully bridge the gap 207 betweenthe piezoelectric elements 12 and the wells 210, then the output of thesignal generator 106 will not be enabled. For output of the signalgenerator 106 to be enabled, the following three conditions must be met:(i) safety interlock state active (that is, the lid 198 is closed); (ii)temperature of transducer fluid 117 is below a threshold temperature;and (iii) level of transducer fluid 117 in the gap 207 is above minimumlevel. However and notwithstanding the above, it will be appreciatedthat in some embodiments it may be desirable to allow testing to makemeasurements with the lid 198 open. In such embodiments, an engineeringmode can permit operation with the lid 198 open.

In various embodiments of the system 100, waveform timing may result insimultaneous and/or sequential driving pulses 108 that do not energizeadjacent piezoelectric elements 12 in the array 112, thereby allowing apiezoelectric element 12 (and adjacent piezoelectric elements 12) tocool down for a period of time before that same piezoelectric element 12is energized again. Waveform timing for embodiments of the system 100will be discussed by way of a non-limiting example for an embodiment ofthe system 100 with a ninety-six well microplate 176, four lenses 16overlie each piezoelectric element 12, and the array 112 includestwenty-four piezoelectric element array assemblies 10. However, it willbe appreciated that similar waveform timing may be applied to anyembodiment of the system 100 in which more than one lens 16 overlies asingle piezoelectric element 12.

Given by way of non-limiting example and referring additionally to FIG.8A, the acoustic waveform that performs the shearing operation (that is,the acoustic waveform of the amplified driving pulses 114) is highamplitude and highly non-linear, as the shearing is primarily a functionof cavitation. The basic waveform timing is shown in FIG. 8A.

Referring additionally to FIG. 8B, the pulsed acoustic waveform isapplied to a column of wells 210 for N Pulses. Then, the column of wells210 is allowed to cool before another burst is applied to the column.The Burst length is equal to an integer number of contiguous Pulseperiods with output acoustic power. Architecturally, the Burst period isequal to six times the Burst length, since there are two of the twelvecolumns being processed at a given time. The overall processing time isan integer multiple of the Burst period.

Referring additionally to FIG. 8C, timing of applied acoustic power tothe respective columns is shown, where t0 through t6 (rows) represent asingle Burst period, and COL1 through COL12 (columns) represent thetwelve columns of the microplate 176. A shaded square 216 indicatesacoustic power output (Burst) and a white square 218 indicates a cooldown period. It will be appreciated that FIG. 8C shows that adjacentpiezoelectric elements 12 are not energized simultaneously (that is, inany single row representing a single Burst period) and are not energizedby sequential waveforms (that is, in any given row that represents agiven Burst period and in a row directly underneath the given row,representing a sequential Burst period).

Referring additionally to FIG. 8D, relative timing of applied acousticpower is shown for the twelve columns where high amplitude 220represents the Burst for a given column. Thus, in FIG. 8D the highamplitudes correspond to the shaded squares of FIG. 8C. As such, FIG. 8Dalso shows that adjacent piezoelectric elements 12 may not be notenergized simultaneously and may not be energized by sequentialwaveforms.

For example and as shown in FIGS. 8C and 8D, it will be appreciated thatthe Burst for COL1 occurs at to. A simultaneous Burst at t0 appliesacoustic power to COL6—which is not adjacent to COL1. Moreover, theBurst at t1 applies acoustic power to COL3 and COL8—neither of which areadjacent to COL1. Further, the soonest that a Burst applies acousticpower to a column adjacent COL1 is at t4—when acoustic power is appliedto COL2. As a result, this timing technique can help to reduce heating.

Referring additionally to FIG. 8E, mapping of the twenty-four channelsof the pulser modules 140 to the piezoelectric elements 12 are shown.Again, it will be appreciated that any number of the pulser modules 140with any number of channels per pulser module 140 may be used asdesired. For example, in some embodiments twenty-four channels of thepulser modules 140 may be implemented with eight pulser modules 140having three channels apiece. However, it will be appreciated that it isnot necessary to process ninety-six samples and any number of samplesmay be processed as desired. Accordingly, fewer than ninety-six wellsmay be processed as desired for a particular application. That is, insome embodiments as few as one well may be processed and in some otherembodiments as many as ninety-six wells may be processed. In suchembodiments, wells that do not contain samples for processing would befilled with water or another liquid that does not contain samples to beprocessed.

Various example embodiments of the disclosed subject matter can bedescribed in view of the following clauses:

1. A piezoelectric element array assembly comprising: at least onepiezoelectric element configured to produce ultrasound energy responsiveto amplified driving pulses; and a lens layer bonded to the at least onepiezoelectric element, the lens layer having a plurality of lensesformed therein that are configured to focus ultrasound energy created bysingle ones of the at least one piezoelectric element into a pluralityof wells of a microplate disposable in ultrasonic communication with thelens layer, wherein more than one of the plurality of lenses overliesingle ones of the at least one piezoelectric element.

2. The piezoelectric element array assembly of Clause 1, wherein the atleast one piezoelectric element includes a column of two piezoelectricelements.

3. The piezoelectric element array assembly of Clause 1, wherein the atleast one piezoelectric element includes a column of four piezoelectricelements.

4. The piezoelectric element array assembly of Clause 1, wherein the atleast one piezoelectric element includes a column of six piezoelectricelements.

5. The piezoelectric element array assembly of Clause 1, wherein the atleast one piezoelectric element includes a column of eight piezoelectricelements.

6. The piezoelectric element array assembly of Clause 1, wherein the atleast one piezoelectric element includes a column of twelvepiezoelectric elements.

7. The piezoelectric element array assembly of Clause 1, wherein fourlenses overlie single ones of the at least one piezoelectric element.

8. The piezoelectric element array assembly of Clause 1, wherein the atleast one piezoelectric element is made of a material including leadzirconate titanate.

9. The piezoelectric element array assembly of Clause 1, wherein thelens layer is made of a material having an acoustic impedance betweenacoustic impedance of the at least one piezoelectric element and acoupling fluid that is disposable between the lens layer and amicroplate.

10. The piezoelectric element array assembly of Clause 1, wherein thelens layer is made of a material chosen from graphite andfluorphlogopite mica in a borosilicate glass matrix.

11. A method of fabricating a piezoelectric element array assembly, themethod comprising: providing at least one piezoelectric elementconfigured to produce ultrasound energy responsive to amplified drivingpulses; and bonding a lens layer to the at least one piezoelectricelement, the lens layer having a plurality of lenses formed therein thatare configured to focus ultrasound energy created by single ones of theat least one piezoelectric element into a plurality of wells of amicroplate disposable in ultrasonic communication with the lens layer,wherein more than one of the plurality of lenses overlie single ones ofthe at least one piezoelectric element.

12. The method of Clause 11, wherein the at least one piezoelectricelement includes a column of two piezoelectric elements.

13. The method of Clause 11, wherein the at least one piezoelectricelement includes a column of four piezoelectric elements.

14. The method of Clause 11, wherein the at least one piezoelectricelement includes a column of six piezoelectric elements.

15. The method of Clause 11, wherein the at least one piezoelectricelement includes a column of eight piezoelectric elements.

16. The method of Clause 11, wherein the at least one piezoelectricelement includes a column of twelve piezoelectric elements.

17. The method of Clause 11, wherein four lenses overlie single ones ofthe at least one piezoelectric element.

18. A system for shearing cellular material, the system comprising: asignal generator configured to generate ultrasound driving pulses; anamplifier electrically coupled to the signal generator and configured toamplify the ultrasound driving pulses; a piezoelectric element arrayincluding at least one piezoelectric element configured to produceultrasound energy responsive to amplified driving pulses; and aplurality of lenses, wherein more than one of the plurality of lensesoverlie single ones of the at least one piezoelectric element andwherein single ones of the plurality of lenses are configured to focusultrasound energy into single ones of a plurality of wells of amicroplate.

19. The system of Clause 18, further comprising: a lens layer bonded tothe at least one piezoelectric element, the lens layer having theplurality of lenses formed therein.

20. The system of Clause 18, wherein single ones of the plurality oflenses are formed in single ones of a plurality of wells in amicroplate.

21. The system of Clause 18, wherein the at least one piezoelectricelement includes a column of two piezoelectric elements.

22. The system of Clause 18, wherein the at least one piezoelectricelement includes a column of four piezoelectric elements.

23. The system of Clause 18, wherein the at least one piezoelectricelement includes a column of six piezoelectric elements.

24. The system of Clause 18, wherein the at least one piezoelectricelement includes a column of eight piezoelectric elements.

25. The system of Clause 18, wherein the at least one piezoelectricelement includes a column of twelve piezoelectric elements.

26. The system of Clause 18, wherein four lenses overlie single ones ofthe at least one piezoelectric element.

27. The system of Clause 18, wherein the at least one piezoelectricelement is made of a material including lead zirconate titanate.

28. The system of Clause 18, wherein the lens layer is made of amaterial having an acoustic impedance between acoustic impedance of theat least one piezoelectric element and a coupling fluid that isdisposable between the lens layer and a microplate.

29. The system of Clause 18, wherein the lens layer is made of amaterial chosen from graphite and fluorphlogopite mica in a borosilicateglass matrix.

30. A method comprising: generating ultrasound driving pulses;amplifying the ultrasound driving pulses; producing ultrasound energywith at least one piezoelectric element responsive to the amplifieddriving pulses; and focusing the ultrasound energy created by singleones of the at least one piezoelectric element into a plurality of wellsof a microplate by a plurality of lenses, wherein more than one of theplurality of lenses overlie single ones of the at least onepiezoelectric element and wherein single ones of the plurality of lensesare ultrasonically coupled to single ones of the plurality of wells.

31. The method of Clause 30, wherein the plurality of lenses are formedin a lens layer that is bonded to the at least one piezoelectricelement.

32. The method of Clause 30, wherein single ones of the plurality oflenses are formed in single ones of a plurality of wells in amicroplate.

33. The method of Clause 30, wherein four lenses overlie single ones ofthe at least one piezoelectric element.

34. A system for shearing cellular material, the system comprising: acomputer processor configured to generate timing signals; a signalgenerator configured to generate ultrasound driving pulses responsive tothe timing signals; an amplifier electrically coupled to the signalgenerator and configured to amplify the ultrasound driving pulses; aplurality of piezoelectric elements arranged in an array of rows andcolumns and configured to produce ultrasound energy responsive toamplified driving pulses, the timing signals being generated such thatadjacent ones of the plurality of piezoelectric elements are notenergized by at least amplified driving pulses chosen from simultaneousdriving pulses and temporally sequential driving pulses; and a pluralityof lenses, wherein more than one of the plurality of lenses overliesingle ones of the plurality of piezoelectric elements and whereinsingle ones of the plurality of lenses are configured to focusultrasound energy into single ones of a plurality of wells of amicroplate.

35. The system of Clause 34, further comprising: a lens layer bonded tothe plurality of piezoelectric elements, the lens layer having aplurality of lenses formed therein.

36. The system of Clause 34, wherein single ones of the plurality oflenses are formed in single ones of a plurality of wells in amicroplate.

37. The system of Clause 34, wherein the plurality of piezoelectricelements includes a column of two piezoelectric elements.

38. The system of Clause 34, wherein the plurality of piezoelectricelements includes a column of four piezoelectric elements.

39. The system of Clause 34, wherein the plurality of piezoelectricelements includes a column of six piezoelectric elements.

40. The system of Clause 34, wherein the plurality of piezoelectricelements includes a column of eight piezoelectric elements.

41. The system of Clause 34, wherein the plurality of piezoelectricelements includes a column of twelve piezoelectric elements.

42. The system of Clause 34, wherein four lenses overlie single ones ofthe at least one piezoelectric element.

43. The system of Clause 34, wherein the at least one piezoelectricelement is made of a material including lead zirconate titanate.

44. The system of Clause 34, wherein the lens layer is made of amaterial having an acoustic impedance between acoustic impedance of theat least one piezoelectric element and a coupling fluid that isdisposable between the lens layer and a microplate.

45. The system of Clause 34, wherein the lens layer is made of amaterial chosen from graphite and fluorphlogopite mica in a borosilicateglass matrix.

46. A system for shearing cellular material, the system comprising: ahousing; a signal generator disposed in the housing and configured togenerate ultrasound driving pulses; an amplifier disposed in the housingand electrically coupled to the signal generator, the amplifier beingconfigured to amplify the ultrasound driving pulses; a piezoelectricelement array disposed in the housing, the piezoelectric element arrayincluding at least one piezoelectric element configured to produceultrasound energy responsive to amplified driving pulses; a plurality oflenses configured to focus ultrasound energy into a plurality of wellsof a microplate; a fluidics system configured to flow therein atransducer fluid; and a seal disposed on the housing, the seal beingconfigured to receive a microplate in sealing engagement thereon suchthat the piezoelectric element array, the housing, and a microplatereceived in sealing engagement on the seal define a chamber in hydrauliccommunication with the fluidics system and configured to contain thereintransducer fluid.

47. The system of Clause 46, further comprising: a lens layer bonded tothe at least one piezoelectric element, the lens layer having theplurality of lenses formed therein.

48. The system of Clause 46, wherein single ones of the plurality oflenses are formed in single ones of a plurality of wells in amicroplate.

49. The system of Clause 46, further comprising: a clamping mechanismconfigured to hold a microplate in sealing engagement on the seal.

50. The system of Clause 46, further comprising: an openably closablelid disposed on the housing.

51. The system of Clause 50, further comprising: an interlock devicemechanically configured to sense position of the lid, the interlockdevice being configured to prevent energization of the piezoelectricelement array when the lid is in an open position.

52. The system of Clause 46, wherein the fluidics system includes areservoir disposed in the housing and configured to receive thereintransducer fluid.

53. The system of Clause 52, wherein the fluidics system furtherincludes a pump disposed in the housing and configured to cause flow oftransducer fluid.

54. The system of Clause 46, wherein the fluidics system includes adebubbling and degassing subsystem.

55. A method of shearing cellular material, the method comprising:placing a microplate with cellular material disposed in a plurality ofwells defined therein on a seal disposed on a housing; clamping themicroplate on the seal in sealing engagement therewith; flowingtransducer fluid in a fluidics system disposed in the housing such thattransducer fluid is placed in hydraulic communication with a pluralityof lenses; energizing an array of piezoelectric elements to produceultrasound energy; and focusing ultrasound energy in the plurality ofwells with a plurality of lenses such that cavitation is induced in thecellular material disposed in the plurality of wells.

56. The method of Clause 55, wherein the plurality of lenses are formedin a lens layer that is bonded to the array of piezoelectric elements.

57. The method of Clause 55, wherein single ones of the plurality oflenses are formed in single ones of the plurality of wells.

58. The method of Clause 55, further comprising: opening a lid in thehousing before placing the microplate with cellular material disposed inthe plurality of wells defined therein on the seal disposed on thehousing; and shutting the lid after clamping the microplate on the sealin sealing engagement therewith.

59. The method of Clause 58, further comprising: satisfying an interlockcondition that permits causing transducer fluid to flow in the fluidicssystem and that permits energizing the array of piezoelectric elementsresponsive to shutting the lid after clamping the microplate on the sealin sealing engagement therewith.

60. The method of Clause 55, wherein: more than one of the plurality oflenses overlie single ones of the plurality of piezoelectric element;and single ones of the plurality of lenses are configured to focusultrasound energy into single ones of the plurality of wells of themicroplate.

61. The method of Clause 60, wherein four lenses overlie single ones ofthe plurality of piezoelectric elements.

62. The method of Clause 55, wherein: the plurality of piezoelectricelements are arranged in an array of rows and columns and are energizedby timed amplified driving pulses; and adjacent ones of the plurality ofpiezoelectric elements are not energized by at least amplified drivingpulses chosen from simultaneous driving pulses and temporally sequentialdriving pulses.

From the foregoing, it will be appreciated that specific embodiments ofthe present subject matter have been described herein for purposes ofillustration, but that various modifications may be made withoutdeviating from the spirit and scope of the present subject matter.Accordingly, the present subject matter is not limited except as by theappended claims.

1. A piezoelectric element array assembly comprising: at least onepiezoelectric element configured to produce ultrasound energy responsiveto amplified driving pulses; and a lens layer bonded to the at least onepiezoelectric element, the lens layer having a plurality of lensesformed therein that are configured to focus ultrasound energy created bysingle ones of the at least one piezoelectric element into a pluralityof wells of a microplate disposable in ultrasonic communication with thelens layer, wherein more than one of the plurality of lenses overliesingle ones of the at least one piezoelectric element. 2-6. (canceled)7. The piezoelectric element array assembly of claim 1, wherein fourlenses overlie single ones of the at least one piezoelectric element. 8.The piezoelectric element array assembly of claim 1, wherein the atleast one piezoelectric element is made of a material including leadzirconate titanate.
 9. The piezoelectric element array assembly of claim1, wherein the lens layer is made of a material having an acousticimpedance between acoustic impedance of the at least one piezoelectricelement and a coupling fluid that is disposable between the lens layerand a microplate. 10-17. (canceled)
 18. A system for shearing cellularmaterial, the system comprising: a signal generator configured togenerate ultrasound driving pulses; an amplifier electrically coupled tothe signal generator and configured to amplify the ultrasound drivingpulses; a piezoelectric element array including at least onepiezoelectric element configured to produce ultrasound energy responsiveto amplified driving pulses; and a plurality of lenses, wherein morethan one of the plurality of lenses overlie single ones of the at leastone piezoelectric element and wherein single ones of the plurality oflenses are configured to focus ultrasound energy into single ones of aplurality of wells of a microplate.
 19. The system of claim 18, furthercomprising: a lens layer bonded to the at least one piezoelectricelement, the lens layer having the plurality of lenses formed therein.20. The system of claim 18, wherein single ones of the plurality oflenses are formed in single ones of a plurality of wells in amicroplate. 21-45. (canceled)
 46. A system for shearing cellularmaterial, the system comprising: a housing; a signal generator disposedin the housing and configured to generate ultrasound driving pulses; anamplifier disposed in the housing and electrically coupled to the signalgenerator, the amplifier being configured to amplify the ultrasounddriving pulses; a piezoelectric element array disposed in the housing,the piezoelectric element array including at least one piezoelectricelement configured to produce ultrasound energy responsive to amplifieddriving pulses; a plurality of lenses configured to focus ultrasoundenergy into a plurality of wells of a microplate; a fluidics systemconfigured to flow therein a transducer fluid; and a seal disposed onthe housing, the seal being configured to receive a microplate insealing engagement thereon such that the piezoelectric element array,the housing, and a microplate received in sealing engagement on the sealdefine a chamber in hydraulic communication with the fluidics system andconfigured to contain therein transducer fluid.
 47. The system of claim46, further comprising: a lens layer bonded to the at least onepiezoelectric element, the lens layer having the plurality of lensesformed therein.
 48. The system of claim 46, wherein single ones of theplurality of lenses are formed in single ones of a plurality of wells ina microplate.
 49. The system of claim 46, further comprising: a clampingmechanism configured to hold a microplate in sealing engagement on theseal.
 50. The system of claim 46, further comprising: an openablyclosable lid disposed on the housing.
 51. The system of claim 50,further comprising: an interlock device mechanically configured to senseposition of the lid, the interlock device being configured to preventenergization of the piezoelectric element array when the lid is in anopen position.
 52. The system of claim 46, wherein the fluidics systemincludes a reservoir disposed in the housing and configured to receivetherein transducer fluid.
 53. The system of claim 52, wherein thefluidics system further includes a pump disposed in the housing andconfigured to cause flow of transducer fluid.
 54. The system of claim46, wherein the fluidics system includes a debubbling and degassingsubsystem.
 55. A method of shearing cellular material, the methodcomprising: placing a microplate with cellular material disposed in aplurality of wells defined therein on a seal disposed on a housing;clamping the microplate on the seal in sealing engagement therewith;flowing transducer fluid in a fluidics system disposed in the housingsuch that transducer fluid is placed in hydraulic communication with aplurality of lenses; energizing an array of piezoelectric elements toproduce ultrasound energy; and focusing ultrasound energy in theplurality of wells with a plurality of lenses such that cavitation isinduced in the cellular material disposed in the plurality of wells. 56.The method of claim 55, wherein the plurality of lenses are formed in alens layer that is bonded to the array of piezoelectric elements. 57.The method of claim 55, wherein single ones of the plurality of lensesare formed in single ones of the plurality of wells.
 58. The method ofclaim 55, further comprising: opening a lid in the housing beforeplacing the microplate with cellular material disposed in the pluralityof wells defined therein on the seal disposed on the housing; andshutting the lid after clamping the microplate on the seal in sealingengagement therewith.
 59. The method of claim 58, further comprising:satisfying an interlock condition that permits causing transducer fluidto flow in the fluidics system and that permits energizing the array ofpiezoelectric elements responsive to shutting the lid after clamping themicroplate on the seal in sealing engagement therewith.
 60. The methodof claim 55, wherein: more than one of the plurality of lenses overliesingle ones of the plurality of piezoelectric element; and single onesof the plurality of lenses are configured to focus ultrasound energyinto single ones of the plurality of wells of the microplate. 61.(canceled)
 62. (canceled)